rss
Br J Ophthalmol 2005;89:658-661 doi:10.1136/bjo.2004.061754
  • Clinical science
    • Scientific reports

In vitro adenovirus mediated gene transfer to the human cornea

  1. C F Jessup,
  2. H M Brereton,
  3. D J Coster,
  4. K A Williams
  1. Department of Ophthalmology, Flinders University, Adelaide, Australia
  1. Correspondence to: Dr Keryn Williams Department of Ophthalmology, Flinders Medical Centre, Bedford Park, South Australia 5042, Australia; keryn.williamsflinders.edu.au
  • Accepted 1 January 2005

Abstract

Background/aims: Replication deficient adenovirus is an efficient vector for gene transfer to the cornea. The aim was to optimise the transduction of human corneal endothelium with adenoviral vectors and to measure transgene production from transduced corneas.

Methods: Adenoviral vectors (AdV) encoding enhanced green fluorescent protein (eGFP) or a transgenic protein (scFv) were used to transfect 34 human corneas. Reporter gene expression was assessed after 72–96 hours of organ culture. The kinetics of scFv production was monitored in vitro for 1 month by flow cytometric analysis of corneal supernatants.

Results: Transduction of human corneas with high doses (5×107–3×108 pfu) of AdV caused eGFP expression in 12–100% of corneal endothelial cells. Corneas were efficiently transduced following up to 28 days in cold storage. Very high AdV doses (2×109 pfu) reduced endothelial cell densities to 98 (SD 129) nuclei/mm2 (compared to 2114 (716) nuclei/mm2 for all other groups). Transgenic protein production peaked at 2.4 (0.9) μg/cornea/day at 2 weeks post-transduction, and decreased to 1.2 (0.4) μg/cornea/day by 33 days, at which time endothelial cell density had decreased to 431 (685) nuclei/mm2.

Conclusion: Human corneas can be efficiently transduced by AdV following extended periods of cold storage, and transgene expression is maintained for at least 1 month in vitro.

Footnotes

  • Funding: This study was supported by the National Health and Medical Research Council of Australia and the Ophthalmic Research Institute of Australia. CFJ is supported by the South Australian Joyner scholarship in Medicine. The funding bodies had no direct input into study design, collection, analysis or interpretation of data for this publication.

  • Competing interests: none declared.

  • Ethics approval: Necessary ethics committee approval was secured for the study from the Flinders Clinical Research Ethics Committee.

This Article

  1. Abstract
  2. Full text
  3. PDF

Responses

  1. Submit a response
  2. No responses published

Register for free content

The full back archive is now available for all BMJ Journals. Institutional subscribers may access the entire archive as part of their subscription. Personal subscribers will also have access to all content when logged in. Non-subscribers who register have free access to all articles published before 2006 right back to volume 1 issue 1. Register here to access the free archive of all BMJ Journals.

Don't forget to sign up for content alerts so you keep up to date with all the articles as they are published.